SMC-5/6 complex subunit NSE-1 plays a crucial role in meiosis and DNA repair in Caenorhabditis elegans.

The SMC5/6 complex is evolutionarily conserved across all eukaryotes and plays a pivotal role in preserving genomic stability. Mutations in genes encoding SMC5/6 complex subunits have been associated with human lung disease, immunodeficiency, and chromosome breakage syndrome. Despite its critical importance, much about the SMC5/6 complex remains to be elucidated. Various evidences have suggested possible role of a subunit of the SMC5/6 complex, NSE1, in chromosome segregation and DNA repair. Current knowledge regarding the role of NSE1 is primarily derived from single-cell-based analyses in yeasts, Arabidopsis thaliana, and human cell lines. However, our understanding of its function is still limited and requires further investigation. This study delves into the role of nse-1 in Caenorhabditis elegans, revealing its involvement in meiotic recombination and DNA repair. nse-1 mutants display reduced fertility, increased male incidence, and increased sensitivity to genotoxic chemicals due to defects in meiotic chromosome segregation and DNA repair. These defects manifest as increased accumulation of RAD-51 foci, increased chromosome fragmentation, and susceptibility to MMS, cisplatin, and HU. Furthermore, nse-1 mutation exacerbates germ cell death by upregulating ced-13 and egl-1 genes involved in the CEP-1/p53-mediated apoptotic pathway. NSE-1 is essential for the proper localization of NSE-4 and MAGE-1 on the chromosomes. Collectively, these findings firmly establish nse-1 as a crucial factor in maintaining genomic stability.

Dual tagging of GFP and Degron on endogenous COSA-1 in Caenorhabditis elegans as a crossover investigation tool.

COSA-1 is essential for accurate meiosis in C. elegans . Two null mutants ( cosa-1 ( me13 ) and cosa-1 ( tm3298 ) ) have been notably studied. These null mutants exhibit severe meiotic defects, hindering the observation of the subtle or dynamic nature of COSA-1 function. To overcome these limitations, we developed a C. elegans strain with inducible COSA-1 degradation using the Auxin-Inducible Degron (AID) system. This strain exhibits normal fertility and COSA-1::GFP foci. Auxin treatment successfully depletes COSA-1, resulting in a 96% decrease in progeny viability and 12 univalent chromosomes in diakinesis oocytes. This strain serves as a valuable tool for studying the dynamics of COSA-1.

Unveiling the super tolerance of Candida nivariensis to oxidative stress: insights into the involvement of a catalase.

Yeast cells involved in fermentation processes face various stressors that disrupt redox homeostasis and cause cellular damage, making the study of oxidative stress mechanisms crucial. In this investigation, we isolated a resilient yeast strain, Candida nivariensis GXAS-CN, capable of thriving in the presence of high concentrations of H2O2. Transcriptomic analysis revealed the up-regulation of multiple antioxidant genes in response to oxidative stress. Deletion of the catalase gene Cncat significantly impacted H2O2-induced oxidative stress. Enzymatic analysis of recombinant CnCat highlighted its highly efficient catalase activity and its essential role in mitigating H2O2. Furthermore, over-expression of CnCat in Saccharomyces cerevisiae improved oxidative resistance by reducing intracellular ROS accumulation. The presence of multiple stress-responsive transcription factor binding sites at the promoters of antioxidative genes indicates their regulation by different transcription factors. These findings demonstrate the potential of utilizing the remarkably tolerant C. nivariensis GXAS-CN or enhancing the resistance of S. cerevisiae to improve the efficiency and cost-effectiveness of industrial fermentation processes.IMPORTANCEEnduring oxidative stress is a crucial trait for fermentation strains. The importance of this research is its capacity to advance industrial fermentation processes. Through an in-depth examination of the mechanisms behind the remarkable H2O2 resistance in Candida nivariensis GXAS-CN and the successful genetic manipulation of this strain, we open the door to harnessing the potential of the catalase CnCat for enhancing the oxidative stress resistance and performance of yeast strains. This pioneering achievement creates avenues for fine-tuning yeast strains for precise industrial applications, ultimately leading to more efficient and cost-effective biotechnological processes.

Ganoderma lucidum methyl ganoderate E extends lifespan and modulates aging-related indicators in Caenorhabditis elegans.

Methyl Ganoderate E (MGE) is a triterpenoid derived from Ganoderma lucidum (Reishi), an edible mushroom, commonly processed into food forms such as soups, drinks, culinary dishes, and supplements. MGE has been shown to inhibit 3T3-L1 murine adipocyte differentiation when combined with other G. lucidum triterpenes. However, the specific effect of MGE on biological processes remains unknown. In this study, we present the first evidence of MGE's anti-aging effect in Caenorhabditis elegans. Through our screening process using the UPRER regulation ability, we evaluated a library of 74 pure compounds isolated from G. lucidum, and MGE exhibited the most promising results. Subsequent experiments demonstrated that MGE extended the lifespan by 26% at 10 µg ml-1 through daf-16, hsf-1, and skn-1-dependent pathways. MGE also enhanced resistance to various molecular stressors, improved healthspan, increased fertility, and reduced the aggregation of alpha-synuclein and amyloid-beta. Transcriptome data revealed that MGE promoted processes associated with proteolysis and neural activity, while not promoting cell death processes. Collectively, our findings suggest that G. lucidum MGE could be considered as a potential anti-aging intervention, adding to the growing list of such interventions.

Caenorhabditis elegans NSE3 homolog (MAGE-1) is involved in genome stability and acts in inter-sister recombination during meiosis.

Melanoma antigen (MAGE) genes encode for a family of proteins that share a common MAGE homology domain. These genes are conserved in eukaryotes and have been linked to a variety of cellular and developmental processes including ubiquitination and oncogenesis in cancer. Current knowledge on the MAGE family of proteins mainly comes from the analysis of yeast and human cell lines, and their functions have not been reported at an organismal level in animals. Caenorhabditis elegans only encodes 1 known MAGE gene member, mage-1 (NSE3 in yeast), forming part of the SMC-5/6 complex. Here, we characterize the role of mage-1/nse-3 in mitosis and meiosis in C. elegans. mage-1/nse-3 has a role in inter-sister recombination repair during meiotic recombination and for preserving chromosomal integrity upon treatment with a variety of DNA-damaging agents. MAGE-1 directly interacts with NSE-1 and NSE-4. In contrast to smc-5, smc-6, and nse-4 mutants which cause the loss of NSE-1 nuclear localization and strong cytoplasmic accumulation, mage-1/nse-3 mutants have a reduced level of NSE-1::GFP, remnant NSE-1::GFP being partially nuclear but largely cytoplasmic. Our data suggest that MAGE-1 is essential for NSE-1 stability and the proper functioning of the SMC-5/6 complex.

Polysaccharides Extracted from Dendrobium officinale Grown in Different Environments Elicit Varying Health Benefits in Caenorhabditis elegans.

Dendrobium officinale is one of the most widely used medicinal herbs, especially in Asia. In recent times, the polysaccharide content of D. officinale has garnered attention due to the numerous reports of its medicinal properties, such as anticancer, antioxidant, anti-diabetic, hepatoprotective, neuroprotective, and anti-aging activities. However, few reports of its anti-aging potential are available. Due to high demand, the wild D. officinale is scarce; hence, alternative cultivation methods are being employed. In this study, we used the Caenorhabditis elegans model to investigate the anti-aging potential of polysaccharides extracted from D. officinale (DOP) grown in three different environments; tree (TR), greenhouse (GH), and rock (RK). Our findings showed that at 1000 µg/mL, GH-DOP optimally extended the mean lifespan by 14% and the maximum lifespan by 25% (p < 0.0001). TR-DOP and RK-DOP did not extend their lifespan at any of the concentrations tested. We further showed that 2000 µg/mL TR-DOP, GH-DOP, or RK-DOP all enhanced resistance to H2O2-induced stress (p > 0.05, p < 0.01, and p < 0.01, respectively). In contrast, only RK-DOP exhibited resistance (p < 0.01) to thermal stress. Overall, DOP from the three sources all increased HSP-4::GFP levels, indicating a boost in the ability of the worms to respond to ER-related stress. Similarly, DOP from all three sources decreased α-synuclein aggregation; however, only GH-DOP delayed ß-amyloid-induced paralysis (p < 0.0001). Our findings provide useful information on the health benefits of DOP and also provide clues on the best practices for cultivating D. officinale for maximum medicinal applications.

An endogenous mCherry-tagged COSA-1 as a crossover investigation tool in Caenorhabditis elegans.

The C. elegans cosa-1 gene encodes the crossover site-associated-1 (COSA-1) protein, a cyclin-related protein that functions in promoting crossovers (COs) during meiosis. Previous studies regarding CO dynamics in live C. elegans have mostly relied on the green fluorescent protein-tagged cosa-1 transgenic strain, which was generated by the microparticle bombardment method. Here, we insert the red fluorescence protein mCherry at the C-terminal of the cosa-1 gene to establish cosa-1::mCherry transgenic worm by the CRISPR/Cas9 technique. The COSA-1::mCherry was observed to appear from the early pachytene, and disappear in the diplotene zone of the germline, with 6 COSA-1:: mCherry foci in the late pachytene, which colocalized with GFP::COSA-1 from AV630 strain. Furthermore, the transgenic strain harboring a cosa-1::mCherry fusion shows no defect in the brood size, progeny viability and male frequency, which provides a useful tool for the meiotic analysis in C. elegans .

Loss of NSE-4 Perturbs Genome Stability and DNA Repair in Caenorhabditis elegans.

The Structural Maintenance of Chromosomes (SMC) complex plays an important role in maintaining chromosome integrity, in which the SMC5/6 complex occupies a central position by facilitating mitotic and meiotic processes as well as DNA repair. NSE-4 Kleisin is critical for both the organization and function of the SMC5/6 complex, bridging NSE1 and NSE3 (MAGE related) with the head domains of the SMC5 and SMC6 proteins. Despite the conservation in protein sequence, no functional relevance of the NSE-4 homologous protein (NSE-4) in Caenorhabditis elegans has been reported. Here, we demonstrated the essential role of C. elegans NSE-4 in genome maintenance and DNA repair. Our results showed that NSE-4 is essential for the maintenance of chromosomal structure and repair of a range of chemically induced DNA damage. Furthermore, NSE-4 is involved in inter-sister repair during meiosis. NSE-4 localizes on the chromosome and is indispensable for the localization of NSE-1. Collectively, our data from this study provide further insight into the evolutionary conservation and diversification of NSE-4 function in the SMC-5/6 complex.

Bioactive Phytochemicals with Anti-Aging and Lifespan Extending Potentials in Caenorhabditis elegans.

In the forms of either herbs or functional foods, plants and their products have attracted medicinal, culinary, and nutraceutical applications due to their abundance in bioactive phytochemicals. Human beings and other animals have employed those bioactive phytochemicals to improve health quality based on their broad potentials as antioxidant, anti-microbial, anti-carcinogenic, anti-inflammatory, neuroprotective, and anti-aging effects, amongst others. For the past decade and half, efforts to discover bioactive phytochemicals both in pure and crude forms have been intensified using the Caenorhabditis elegans aging model, in which various metabolic pathways in humans are highly conserved. In this review, we summarized the aging and longevity pathways that are common to C. elegans and humans and collated some of the bioactive phytochemicals with health benefits and lifespan extending effects that have been studied in C. elegans. This simple animal model is not only a perfect system for discovering bioactive compounds but is also a research shortcut for elucidating the amelioration mechanisms of aging risk factors and associated diseases.

Effects of various inhibitory substances and immobilization on ethanol production efficiency of a thermotolerant Pichia kudriavzevii.

BACKGROUND: Although bioethanol production has been gaining worldwide attention as an alternative to fossil fuel, ethanol productivities and yields are still limited due to the susceptibility of fermentation microorganisms to various stress and inhibitory substances. There is therefore an unmet need to search for multi-stress-tolerant organisms to improve ethanol productivity and reduce production cost, particularly when lignocellulosic hydrolysates are used as the feedstock. RESULTS: Here, we have characterized a previously isolated Pichia kudriavzevii LC375240 strain which is thermotolerant to high temperatures of 37 °C and 42 °C. More excitingly, growth and ethanol productivity of this strain exhibit strong tolerance to multiple stresses such as acetic acid, furfural, formic acid, H2O2 and high concentration of ethanol at 42 °C. In addition, simple immobilization of LC375240 on corncobs resulted to a more stable and higher efficient ethanol production for successive four cycles of repeated batch fermentation at 42 °C. CONCLUSION: The feature of being thermotolerant and multi-stress-tolerant is unique to P. kudriavzevii LC375240 and makes it a good candidate for second-generation bioethanol fermentation as well as for investigating the molecular basis underlying the robust stress tolerance. Immobilization of P. kudriavzevii LC375240 on corncobs is another option for cheap and high ethanol productivity.

Identification of a Novel Transcription Factor TP05746 Involved in Regulating the Production of Plant-Biomass-Degrading Enzymes in Talaromyces pinophilus.

Limited information on transcription factor (TF)-mediated regulation exists for most filamentous fungi, specifically for regulation of the production of plant-biomass-degrading enzymes (PBDEs). The filamentous fungus, Talaromyces pinophilus, can secrete integrative cellulolytic and amylolytic enzymes, suggesting a promising application in biotechnology. In the present study, the regulatory roles of a Zn2Cys6 protein, TP05746, were investigated in T. pinophilus through the use of biochemical, microbiological and omics techniques. Deletion of the gene TP05746 in T. pinophilus led to a 149.6% increase in soluble-starch-degrading enzyme (SSDE) production at day one of soluble starch induction but an approximately 30% decrease at days 2 to 4 compared with the parental strain ΔTpKu70. In contrast, the T. pinophilus mutant ΔTP05746 exhibited a 136.8-240.0% increase in raw-starch-degrading enzyme (RSDE) production, as well as a 90.3 to 519.1% increase in cellulase and xylanase production following induction by culturing on wheat bran plus Avicel, relative to that exhibited by ΔTpKu70. Additionally, the mutant ΔTP05746 exhibited accelerated mycelial growth at the early stage of cultivation and decreased conidiation. Transcriptomic profiling and real-time quantitative reverse transcription-PCR (RT-qPCR) analyses revealed that TP05746 dynamically regulated the expression of genes encoding major PBDEs and their regulatory genes, as well as fungal development-regulated genes. Furthermore, in vitro binding experiments confirmed that TP05746 bound to the promoter regions of the genes described above. These results will contribute to our understanding of the regulatory mechanism of PBDE genes and provide a promising target for genetic engineering for improved PBDE production in filamentous fungi.

The transcription factor TpRfx1 is an essential regulator of amylase and cellulase gene expression in Talaromyces pinophilus.

BACKGROUND: Perfect and low cost of fungal amylolytic and cellulolytic enzymes are prerequisite for the industrialization of plant biomass biorefinergy to biofuels. Genetic engineering of fungal strains based on regulatory network of transcriptional factors (TFs) and their targets is an efficient strategy to achieve the above described aim. Talaromyces pinophilus produces integrative amylolytic and cellulolytic enzymes; however, the regulatory mechanism associated with the expression of amylase and cellulase genes in T. pinophilus remains unclear. In this study, we screened for and identified novel TFs regulating amylase and/or cellulase gene expression in T. pinophilus 1-95 through comparative transcriptomic and genetic analyses. RESULTS: Comparative analysis of the transcriptomes from T. pinophilus 1-95 grown on media in the presence and absence of glucose or soluble starch as the sole carbon source screened 33 candidate TF-encoding genes that regulate amylase gene expression. Thirty of the 33 genes were successfully knocked out in the parental strain T. pinophilus ∆TpKu70, with seven of the deletion mutants firstly displaying significant changes in amylase production as compared with the parental strain. Among these, ∆TpRfx1 (TpRfx1: Talaromyces pinophilus Rfx1) showed the most significant decrease (81.5%) in amylase production, as well as a 57.7% reduction in filter paper cellulase production. Real-time quantitative reverse transcription PCR showed that TpRfx1 dynamically regulated the expression of major amylase and cellulase genes during cell growth, and in vitro electrophoretic mobility shift assay revealed that TpRfx1 bound the promoter regions of genes encoding α-amylase (TP04014/Amy13A), glucoamylase (TP09267/Amy15A), cellobiohydrolase (TP09412/cbh1), ß-glucosidase (TP05820/bgl1), and endo-ß-1,4-glucanase (TP08514/eg1). TpRfx1 protein containing a regulatory factor X (RFX) DNA-binding domain belongs to RFX family. CONCLUSION: We identified a novel RFX protein TpRFX1 that directly regulates the expression of amylase and cellulase genes in T. pinophilus, which provides new insights into the regulatory mechanism of fungal amylase and cellulase gene expression.

Deletion of TpKu70 facilitates gene targeting in Talaromyces pinophilus and identification of TpAmyR involvement in amylase production.

Talaromyces pinophilus is a promising filamentous fungus for industrial production of biomass-degrading enzymes used in biorefining, and its genome was recently sequenced and reported. However, functional analysis of genes in T. pinophilus is rather limited owing to lack of genetic tools. In this study, a putative TpKu70 encoding the Ku70 homolog involved in the classic non-homologous end-joining pathway was deleted in T. pinophilus 1-95. ΔTpKu70 displayed no apparent defect in vegetative growth and enzyme production, and presented similar sensitivity to benomyl, bleomycin, and UV, when compared with the wild-type T. pinophilus strain 1-95. Seven genes that encode putative transcription factors, including TpAmyR, were successfully knocked out in ΔTpKu70 at 61.5-100% of homologous recombination frequency, which is significantly higher than that noted in the wild-type. Interestingly, ΔTpAmyR produced approximately 20% of amylase secreted by the parent strain ΔTpKu70 in medium containing soluble starch from corn as the sole carbon source. Real-time quantitative reverse transcription PCR showed that TpAmyR positively regulated the expression of genes encoding α-amylase and glucoamylase. Thus, this study provides a useful tool for genetic analysis of T. pinophilus, and identification of a key role for the transcription factor TpAmyR in amylase production in T. pinophilus.